PROJECT SUMMARY/ ABSTRACT Despite effective antiretroviral therapy (ART), HIV-infected individuals have reduced life expectancy and a higher incidence of aging-associated diseases compared to HIV-uninfected controls. Persistent systemic inflammation despite suppressive ART has been associated with serious non-AIDS morbidity (e.g., myocardial infarction, stroke, malignancy) and mortality. Recent data suggests that an upstream regulator of interleukin (IL)-6, interleukin-1 beta (IL-1?), may be the major driver of increased cardiovascular risk observed in HIV+ ART-suppressed individuals. The recent CANTOS trial has now demonstrated in over 10,000 individuals that in vivo IL-1? blockade with the monoclonal antibody canakinumab significantly reduced cardiovascular events and cancer mortality in the general population. We have recently performed a phase 1 trial administering a single dose of canakinumab to HIV+ ART-suppressed participants and found that in vivo IL-1? blockade led to significant reductions in plasma IL-1? (as well as associated systemic inflammatory markers plasma IL-6 and high sensitivity C-reactive protein), vascular inflammation, and monocyte activation. Furthermore, our ex vivo data suggests that IL- ? plays a critical role in maintaining the ?HIV reservoir? (the total amount of residual virus that persists during ART suppression and potentially drives systemic inflammation). We will use an unbiased integrated approach that combines several high dimensional datasets to test the hypothesis that IL-1? triggers the proinflammatory response that fuels HIV immune dysfunction and persistence. Our proposed study will be the first (in HIV+ or HIV-uninfected individuals) to pursue the link between host genetics and plasma IL-1? levels ? while simultaneously assaying several proinflammatory cytokines in this pathway, including IL-6 and IL-18. In Aim 1 we will identify DNA variants associated with plasma IL-1? in 1,000 HIV+ ART-suppressed participants from the Center for AIDS Research Network of Integrated Clinical Systems (CNICS) cohort using custom whole exome sequencing to add to existing genomewide array data. Individuals with extreme phenotypes (highest and lowest deciles of plasma IL-1? levels) will then be selected for functional validation in Aim 2 using a novel approach that simultaneously characterizes RNA and protein expression at single cell resolution using single cell RNA and antibody sequency (scRNA-Abseq). Finally, we will perform functional in vivo validation of identified genes associated with IL-1? signaling, leveraging samples from our phase 1 trial of canakinumab in Aim 3. Therefore, we will functionally validate findings from Aim 1 as well as identify novel genes/pathways by studying individuals with unique HIV+ phenotypes (extreme plasma IL-1? levels and after canakinumab treatment, respectively) in Aims 2 and 3. The proposed work will identify specific genes and immune pathways that may act synergistically with IL-1?, identifying novel therapeutic targets that may be broadly applicable for the treatment of inflammation-associated diseases in HIV+ and non-HIV populations.